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The mechanism of biosynthesis and direction of chain extension of a poly-(N-acetylglucosamine 1-phosphate) from the walls of Staphylococcus lactis N.C.T.C. 2102.

ABSTRACT: 1. The synthesis of a polymer of N-acetylglucosamine 1-phosphate, occurring in the walls of Staphylococcus lactis N.C.T.C. 2102, was examined by using cell-free enzyme preparations. The enzyme system was particulate, and probably represents fragmented cytoplasmic membrane. 2. Uridine diphosphate N-acetylglucosamine was the only substrate required for polymer synthesis and labelled substrate was used to show that N-acetylglucosamine 1-phosphate is transferred as an intact unit from substrate to polymer. 3. The properties of the enzyme system were studied. A high concentration of Mg(2+) or Mn(2+) was required for optimum activity, and the pH optimum was about 8.5. 4. End-group analysis during synthesis in vitro showed that newly formed chains contain up to about 15 repeating units. Pulse-labelling indicated that chain extension occurs by transfer from the nucleotide to the ;sugar-end' of the chain, i.e. to the end that is not attached to peptidoglycan in the wall.

SUBMITTER: Brooks D

PROVIDER: S-EPMC1184743 | biostudies-other | 1969 Jul

REPOSITORIES: biostudies-other

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Project description:Staphylococcus aureus clinical strains are able to produce at least two distinct types of biofilm matrixes: biofilm matrixes made of the polysaccharide intercellular adhesin (PIA) or poly-N-acetylglucosamine (PNAG), whose synthesis is mediated by the icaADBC locus, and biofilm matrixes built of proteins (polysaccharide independent). σB is a conserved alternative sigma factor that regulates the expression of more than 100 genes in response to changes in environmental conditions. While numerous studies agree that σB is required for polysaccharide-independent biofilms, controversy persists over the role of σB in the regulation of PIA/PNAG-dependent biofilm development. Here, we show that genetically unrelated S. aureus σB-deficient strains produced stronger biofilms under both static and flow conditions and accumulated higher levels of PIA/PNAG exopolysaccharide than their corresponding wild-type strains. The increased accumulation of PIA/PNAG in the σB mutants correlated with a greater accumulation of the IcaC protein showed that it was not due to adjustments in icaADBC operon transcription and/or icaADBC mRNA stability. Overall, our results reveal that in the presence of active σB, the turnover of Ica proteins is accelerated, reducing the synthesis of PIA/PNAG exopolysaccharide and consequently the PIA/PNAG-dependent biofilm formation capacity.IMPORTANCE Due to its multifaceted lifestyle, Staphylococcus aureus needs a complex regulatory network to connect environmental signals with cellular physiology. One particular transcription factor, named σB (SigB), is involved in the general stress response and the expression of virulence factors. For many years, great confusion has existed about the role of σB in the regulation of the biofilm lifestyle in S. aureus Our study demonstrated that σB is not necessary for exopolysaccharide-dependent biofilms and, even more, that S. aureus produces stronger biofilms in the absence of σB The increased accumulation of exopolysaccharide correlates with higher stability of the proteins responsible for its synthesis. The present findings reveal an additional regulatory layer to control biofilm exopolysaccharide synthesis under stress conditions.

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The mechanism of biosynthesis and direction of chain extension of a poly-(N-acetylglucosamine 1-phosphate) from the walls of Staphylococcus lactis N.C.T.C. 2102.

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